Microsphere-based drug delivery systems are advantageous because of their injectability and versatility in controlling the release patterns of the loaded drugs (Sinha and Trehan, J. Control. Release, 90(3):261-80 (2003)). This reduces invasiveness of multiple injections. Biocompatibility and biodegradability are necessary criteria for selecting the drug carriers. Both synthetic polymers such as polylactic acids and polyglycolic acids and natural polymers such as chitosan and alginates can be used for making microspheres for drug delivery (Sinha and Trehan, J. Control. Release, 90(3):261-80 (2003)). Poly-lactic-glycolic acid (PLGA) microspheres dominate this field (Pean, et al., J. Control. Release, 56(1-3):175-87 (1998); Sinha and Trehan, J. Control. Release, 90(3):261-80 (2003); Jiang, et al., Adv. Drug Deliv. Rev., 57(3):391-410 (2005)) because it has been used for years as a suture material. However, as a polyester, PLGA has inevitable intrinsic shortcomings (Sinha and Trehan, J. Control. Release, 90(3):261-80 (2003)) such as low protein polymer compatibility due to the limited solubility and stability of protein in the hydrophobic PLGA matrix, and the extreme acidity of the degradation products at local injury site, which can damage cells and denature proteins. Typically fabricated by a double-emulsion technique, PLGA microspheres often show low encapsulation efficiency and poor retention of bioactivity of the encapsulated protein. Previous reports have demonstrated large burst effect of nerve growth factor (“NGF”) release and loss of bioactivity as early as 48 hours (Pean, et al., J. Control. Release, 56(1-3):175-87 (1998); Hadlock, et al., J. Reconstr. Microsurg., 19(3):179-84; discussion 185-6 (2003)). Apart from instability of the protein drugs, high initial burst and incomplete release also affect the efficiency of microsphere-based drug delivery systems (Yeo and Park, Arch. Pharm. Res., 27(1):1-12 (2004)).
Strategies aiming to improve the initial rapid loss using methods enhancing the protein distribution throughout the polymer matrix (Fu, et al., J. Pharm. Sci., 92(8):1582-91 (2003)) have been reported. Polymerization, emulsification, spray drying and solvent extraction or combinations of these processes are commonly used methods for preparation of polymeric microspheres (Freiberg and Zhu, Int. J. Pharm., 282(1-2):1-18 (2004)). U.S. Pat. No. 6,630,156 to Seo, et al. discloses a method for producing polymer microspheres incorporating physiologically active molecules by emulsification followed by solvent extraction. These methods involve the usage of organic solvents, emulsifying stabilizer and vigorous stirring. These present chemical and mechanical stresses, which can exert damaging effects on the conformational and biological integrity of many drugs in particular proteins (Yeo and Park, Arch. Pharm. Res., 27(1):1-12 (2004)). Moreover, the acidic and hydrophobic microenvironment within the degrading polymers can further damage the loaded drugs (Freiberg and Zhu, Ins. J. Pharm., 282(1-2):1-18 (2004)).
Natural extracellular matrix such as collagen has excellent biocompatibility and negligible immunogenicity (Sano, et al., Adv. Drug Deliv. Rev., 31(3):247-266 (1998); Lee, et al. 2001), and excellent protein compatibility. Therefore, they are excellent candidates for protein delivery devices. These materials provide the natural extracellular milieu that stabilize proteins and potentiate or augment the activity of protein drugs such as growth factors (Lee, et al., Int. J. Pharm., 221(1-2):1-22 (2001); Jones, et al., J. Physiol., 533(1):83-9 (2001); Milev, et al., J. Biol. Chem., 273(34):21439-42 (1998)). Moreover, degradation of these materials results in naturally occurring monomers at neutral pH that do not generate local injury or inflammation. Furthermore, these materials facilitate cell adhesion, attachment and growth that may help in efficiently delivering the signals regulating cell activities. However, their development as drug delivery devices have been overshadowed by advances in synthetic polymers due to the poor dimensional and mechanical stability, and rapid swelling properties of these natural extracellular matrix biomaterials (Yannas, et al. IV. Natural materials. In: Ratner B D, Hoffman A S, Schoen F J, Lemons J E, editors. Biomaterials Sciences—An introduction to materials in medicine. California; Academic Press, 1996: 84-93). This is because most microsphere fabrication method requires vigorous mixing that may fragment these materials (Freiberg and Zhu, Int. J. Pharm., 282(1-2):1-18 (2004)). Therefore, it is almost impossible to fabricate microspheres using these materials unless chemical crosslinking has been used. However, toxicity associated with the residue chemical crosslinking agent prevents its use in drug delivery (Sinha and Trehan, J. Control. Release, 90(3):261-80 (2003)). US Patent Application 20060222680 by Yang and Mark discloses a method of preparing chemically crosslinked collagen microspheres. Chemical crosslinking using glutaraldehyde is efficient in crosslinking the polymers with enhanced mechanical and shape stability. However, it compromises the biocompatibility of the crosslinked structures because the toxic residual chemicals and degradation products induce cytotoxicity and calcification (Simmons, et al., Biotechnol. Appl. Biochem., 17 (Pt 1):23-9 (1993)).
Drug release from a matrix carrier is controlled by either diffusion or degradation or in many cases combinations of two. Many formulation parameters, including the drug, matrix and environmental factors affect the initial burst and the rate of release (Yeo and Park, Arch. Pharm. Res., 27(1):1-12 (2004)). Drug factors such as surface charge, hydrophobicity, loading and solubility in the continuous phase of the drug may affect the interaction between the drug and the matrix and thus the initial burst and release rate. Matrix factors such as the hydrophilicity, concentration, porosity, density, mesh size and swelling properties of the matrix also affect the interaction between the drug and the matrix and thus the initial burst and release rate.
It is therefore an object of the present invention to provide microparticles formed of ECM materials that provide controlled release of bioactive materials and desirable mechanical properties, and methods of manufacture and use thereof.
It is another object of the invention to provide microparticles formed of ECM materials that are highly compatible with bioactive materials and particularly peptides, polypeptides and proteins.
It is a further object of the present invention to provide methods of manufacture that are simple, mild, and non-toxic, without vigorous stirring or organic solvents.